反向遗传技术致弱的基因Ⅶ型新城疫病毒对SPF鸡的免疫效力评价

摘要

为评估致弱的基因Ⅶ型新城疫病-毒(NDV)的免疫效力,本研究采用反向遗传技术构建的一株基因Ⅶ型NDV减毒疫苗株(MG7株),通过滴鼻点眼接种4周龄SPF鸡后,检测其体液免疫、细胞免疫以及攻毒后保护效率和排毒情况等指标,评价MG7株的免疫效果.结果表明:MG7株免疫3周后,能够诱导SPF鸡抗体的产生,且抗体水平(＞6.0810g2)高于LaSota株免疫组(5.210g2).MG7株免疫1～3周内,PBMCs中CD8α+T淋巴细胞亚群比例呈逐渐上升趋势;MG7免疫后第3周,PBMCs中CD4+T淋巴细胞亚群的比例明显增加(p＜0.05);同时,MG7株免疫组的血清IFN-γ含量也呈递增趋势,与空白对照组存在极显著差异(p＜0.01).此外,以基因Ⅶ型NDV强毒株进行攻毒试验,在攻毒后第2d～6d,MG7株免疫组SPF鸡未检测出排毒;LaSota株免疫组则存在不同程度排毒现象,而空白对照组全部死亡.实验表明,与LaSota株免疫组相比,MG7株能够有效诱导NDV特异性抗体产生和T细胞免疫反应,提高SPF鸡的保护效率和有效阻止排毒.本研究对MG7株免疫效力的评估结果表明其具有作为新型基因Ⅶ型NDV疫苗株的潜力,有助于我国ND的防控和根除.%Currently,Newcastle disease virus (NDV) vaccines developed with genotype Ⅰ and Ⅱvirus strains are widely used in practice in China.However,the antigenic difference does exist between vaccine strain and epidemic NDV strain in the field in China,which has resulted in the incomplete protection and virus shedding from chickens,even several sporadic ND outbreaks accidently after vaccination.In the present study,an attenuated NDV (MG7 strain) engineered by the reverse genetics technique in our laboratory was used to evaluate the immune efficacy in SPF chickens via the intranasal and ocular routes.Humoral immunity,cellular immunity,protection efficiency,virus shedding and other indicators were tested to evaluate the immune effect of MG7 strain.The results showed that MG7 strain induced specific antibodies,and the antibody level of MG7 group (＞6.08 log2) was higher than that of the LaSota group (5.2 log2) at three weeks after immunization.The percentage of CD8α+ T cells in PBMCs increased gradually from 1 to 3 weeks post vaccination,and the percentage of CD4+ T cells in PBMCs was significantly elevated at 3 weeks post vaccination (p＜0.05).At the same time,the concentrations of IFN-γ in the chicken sera from MG7 group was increased,which was significantly different from that of control group (p＜0.01).In addition,the virus shedding was detected from SPF chickens in the LaSota group,but not in the MG7 group at 2 to 6 days after challenge,while the chickens in the control group were all dead.Taken together,MG7 strain could effectively induce NDV specific antibody and T cell immune response,improve the protective efficiency and prevent the virus shedding in comparison to LaSota strain.These results suggest MG7 strain has a potential to be a novel vaccine strain candidate against NDV,facilitating ND prevention and eradication.